Background:
Lynch syndrome (also known as Hereditary Non-Polyposis Colon Cancer, HNPCC), is an autosomal dominant hereditary cancer syndrome that accounts for 3-5% of all colon cancers. Lynch syndrome-associated tumors are characterized by microsatellite instability (MSI) and are caused by germline mutations in any of four mismatch repair (MMR) genes (MLH1, MSH6, MSH2, PMS2). The risk of colon and gastric cancers are increased in both sexes, and women with Lynch syndrome have an increased risk for endometrial and ovarian cancers.
The testing strategy for Lynch syndrome includes screening by MSI analysis followed by immunohistochemistry (IHC) testing of MMR proteins. Full gene sequencing can then be performed to identify germline mutations in the putative mutated gene(s) identified by IHC (please contact our lab for more information regarding MSI and IHC testing). Germline mutations in MLH1 and MSH2 account for 90% of Lynch syndrome cases while mutations of MSH6 and PMS2 comprise the remaining fractions. While one study indicated that 65% of MSH2 and 87% of MLH1 variants were point mutations (the rest being gross deletions/duplications), comprehensive clinical sensitivity of MSH2, MLH1, and MSH6 sequencing is unknown.
Reasons for Referral:
- Unknown familial mutation in MMR gene(s) with a positive family history of colorectal cancer without IHC and/or MSI information.
- Identification of inherited genetic defects in MMR gene(s) in colorectal cancer patients with tumors testing positive by IHC and/or MSI.
- Predispositional testing for asymptomatic family members with a positive family history of colorectal cancer.
Methodology:
Sequencing for MLH1, MSH2, MSH6 is carried out by amplification of all exons and intron/exon boundaries followed by bi-directional Sanger sequencing.
This assay does not assess regions of insufficient coverage, introns and promoter regions; pseudogenes; where the reference genome is inaccurate or contains gaps and insertions; and regions of high GC or polynucleotide repeats, but may contain variants that impact gene function. Promoter, intronic, etc. regions are not assessed on our assay, but may contain variants that impact gene function.
Exon-level deletion/duplication analysis is performed by running the NGS data through the Genome Analysis Toolkit (GATK) Germline Copy Number Variation best practices pipeline from GATK, version 4.1.4.1. A Bayesian model was validated clinically in our lab. The model can detect copy changes at a resolution of three (3) or more probe targets (exons) for deletions and duplications in genes that do not have pseudogenes, and is not designed to detect low-level mosaicism or balanced alterations.
Specimen Requirements:
Blood: EDTA or ACD (Solution A or B):
- Adult: 5 mL
- Child: 5 mL
- Infant: 2-3 mL
Saliva: 2 ORAgene™ Assisted Saliva Collection Kits (OGR-500) used according to manufacturer instructions. Please contact KDL Client Services for a Saliva Collection Kit for patients that cannot provide a blood sample.
Assisted Saliva: 4 ORAgene™ Assisted Saliva Collection Kits (OGR-575) used according to manufacturer instructions. Please contact KDL Client Services for an Assisted Saliva Collection Kit for patients that cannot provide a blood sample.
Skin Fibroblast: Punch Biopsy (Cell cultures will be prepared at KDL and used for testing), or 2 T-25 confluent flasks.
DNA: 5-10µg at a minimum of 100ng/µL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS).
A REQUISITION FORM MUST ACCOMPANY ALL SAMPLES. Please include detailed clinical information, including ethnicity, clinical history, and family history.
Test Performed (Days):
Weekly
Turn Around Time:
14-21 days
Shipment Sensitivity Requirements:
- Package and ship specimens to remain cold, but not frozen.
- Ship via overnight express, using the FedEx priority overnight label provided.
- Contact Client Services for shipping kits and instructions at (855) 535-1522.
References:
- Balmaña J, Stockwell DH, Steyerberg EW, et al. Prediction of MLH1 and MSH2 Mutations in Lynch Syndrome. JAMA: The Journal of the American Medical Association. 2006;296(12):1469 -1478.
- Bonadona, V. et al., 2011. Cancer Risks Associated With Germline Mutations in MLH1, MSH2, and MSH6 Genes in Lynch Syndrome. JAMA: The Journal of the American Medical Association, 305(22), pp.2304-2310.
- Evaluation of Genomic Applications in Practice and Prevention (EGAPP) Working Group, 2009. Recommendations from the EGAPP Working Group: genetic testing strategies in newly diagnosed individuals with colorectal cancer aimed at reducing morbidity and mortality from Lynch syndrome in relatives. Genetics in Medicine, 11(1), pp.35-41.
- Goodfellow, P.J. et al., 2003. Prevalence of defective DNA mismatch repair and MSH6 mutation in an unselected series of endometrial cancers. Proceedings of the National Academy of Sciences, 100(10), pp.5908-5913.
Additional Info: